Antibiotic Susceptibility and Virulence Genes in <i>Enterococcus</i> Isolates from Wild Mammals Living in Tuscany, Italy

Abstract

Drug resistance is of great importance to human and animal health, but wild environments are still poorly understood in terms of their function as reservoirs of dangerous microbes and resistance determinants. The aim of the study was to determine the antibiotic susceptibility and virulence factors of <i>Enterococcus</i> bacteria from wildlife in Tuscany, Italy. Of the 36 mammalian fecal samples, 52 isolates were derived and classified as <i>Enterococcus faecium</i> (46% of isolates), <i>Enterococcus hirae</i> (19%), <i>Enterococcus faecalis</i> (13%), <i>Enterococcus gallinarum</i> (8%), <i>Enterococcus casseliflavus</i> (6%), <i>Enterococcus durans</i> (4%), <i>Enterococcus mundtii</i> (2%), and <i>Enterococcus canintestini</i> (2%) using both matrix-assisted laser desorption/ionization-time of flight mass spectrometry and methods based on analysis of genetic material. The isolates tested showed the most frequent resistance to tetracycline (36.5% isolates), ciprofloxacin (36.5%), and erythromycin (25%). Three isolates showed high level of resistance (minimal inhibitory concentration ≥1,024 μg/mL) to vancomycin and teicoplanin, and 15% of the isolates demonstrated multidrug resistance. No isolate resistant to ampicillin, linezolid, or streptomycin was found. Among resistance genes, <i>aac(6)-Ii</i> (50% isolates), <i>msrA/B</i> (48%), <i>msrC</i> (42%), and <i>tetM</i> (31%) were identified most frequently. All <i>E. faecium</i> and <i>E. faecalis</i> isolates were positive for the <i>efaAfm</i> and <i>efaAfs</i> genes, respectively. Other virulence-associated genes, that is, <i>gelE</i>, <i>cylA</i>, <i>asa1</i>, <i>esp</i>, <i>ace</i>, <i>orf1481</i>, <i>ptsD</i>, and <i>sgrA</i>, were found in the majority of <i>E. faecalis</i> and several <i>E. faecium</i> isolates. Multilocus sequence typing analysis performed for selected isolates revealed three new sequence types. The results obtained indicate that wild mammals might act as reservoirs of resistance and virulence determinants that could be transferred between different ecosystems.